Abstract

The mechanism of primary urine production and the activity of a diuretic kinin, Achdo-KII, were investigated in Malpighian tubules of Acheta domesticus by measuring intracellular Na+, K+ and Cl− activities, basolateral membrane voltage (Vb), fluid secretion and transepithelial ion transport. Calculated electrochemical gradients for K+ and Cl− across the basolateral membrane show they are actively transported into principal cells, and basolateral Ba2+-sensitive K+ channels do not contribute to net transepithelial K+ transport and fluid secretion. A basolateral Cl− conductance was revealed after the blockade of K+ channels with Ba2+, and a current carried by the passive outward movement of Cl− accounts for the hyperpolarization of Vb in response to Ba2+. Ion uptake via Na+/K+/2Cl− cotransport, driven by the inwardly directed Na+ electrochemical gradient, is thermodynamically feasible, and is consistent with the actions of bumetanide, which reduces fluid secretion and both Na+ and K+ transport. The Na+ gradient is maintained by its extrusion across the apical membrane and by a basolateral ouabain-sensitive Na+/K+-ATPase. Achdo-KII has no significant effect on the intracellular ion activities or Vb. Electrochemical gradients across the apical membrane were estimated from previously published values for the levels of Na+, K+ and Cl− in the secreted fluid. The electrochemical gradient for Cl− favours passive movement into the lumen, but falls towards zero after stimulation by Achdo-KII. This coincides with a twofold increase in Cl− transport, which is attributed to the opening of an apical Cl− conductance, which depolarises the apical membrane voltage.